The major objective of these studies is to examine cell replication as a function of aging in cultured human cells in vitro and in vivo in rats and mice. To examine the question of whether age-related alterations seen in vitro reflect in vivo cellular aging, cell cultures established from young and old healthy human donors were examined for their in vitro lifespans and cell replicative capabilities. Significant decreases in fibroblast outgrowth, cumulative cell population doublings and cell population replication rates were found in the skin fibroblast cultures derived from the old donor group when compared to the cultures derived from young donors. Cell separation studies based on differences in cell volumes were initiated in an attempt to obtain cell populations that were enriched for either rapidly replicating cells or slow or nonreplicating cells. These studies successfully isolated out populations of senescent cells whose cell volumes and percent of rapidly replicating cells clearly resembled young or early passage cell cultures. However, upon reintroduction of these cells into tissue culture conditions, they rapidly returned to the appearance of senescent cell populations. Further studies are being directed at developing in vivo systems for accurately measuring cell replication.